Project 4 SUMMARY/ABSTRACT Over the last three decades, 5-year survival has improved for patients with epithelial ovarian cancer, but long- term survival has not changed and remains at approximately 30% overall. Following surgical cytoreduction, all new patients receive standard primary chemotherapy that includes a combination of carboplatin and paclitaxel. Primary therapy with platinum-based compounds alone produces regression in approximately 70% of ovarian cancers, whereas 42% respond to paclitaxel alone and there is no synergy between the two drugs. Our long- term translational goal is to increase the effectiveness of chemotherapy for ovarian cancer. While many investigators have focused on overcoming acquired resistance to taxanes, relatively little attention has been given to enhancing paclitaxel response during primary chemotherapy. Using a functional siRNA screen for kinases that regulate sensitivity to paclitaxel, we found that knockdown of the serine-threonine kinase salt- induced kinase 2 (SIK2) induces polyploidy, inhibits ovarian cancer growth and enhances paclitaxel sensitivity in cell lines and xenografts. SIK2 is required for normal centrosome splitting and is overexpressed in 34% of ovarian cancers of all histotypes, associated with decreased overall survival. During mitosis, SIK2 undergoes autophosphorylation and phosphorylates C-Nap1 a structural protein that mediates centrosome splitting. SIK2 also phosphorylates p85?, driving activation of PI3K, as well as HDAC5, modifying chromatin and DNA repair. We have established a collaboration with Arrien Pharmaceuticals to develop orally administered small molecular weight inhibitors of SIK2: ARN-3236 and ARN-3261 that differ by a single solvent binding substitution. ARN-3236 inhibited growth of 10 ovarian cancer cell lines at an IC50 of 0.8 to 2.6 ?M, where the IC50 of ARN-3236 was inversely correlated with endogenous SIK2 expression (Pearson?s r = -0.642, P = 0.03). ARN-3236 also enhanced sensitivity to paclitaxel in 8 of 10 cell lines, as well as in SKOv3ip (P = 0.028) and OVCAR8 xenografts. ARN-3236 uncoupled the centrosome from the nucleus in interphase, blocked centrosome separation in mitosis, caused prometaphase arrest and induced apoptotic cell death as well as tetraploidy. ARN-3236 also inhibited AKT phosphorylation and attenuated survivin expression. ARN-3261 enhanced sensitivity to paclitaxel and cisplatin in xenograft models and exhibited little toxicity, as well as greater resistance to PgP and13-fold greater potency than ARN-3236. We will pursue three aims: 1) to perform a phase I trial of the SIK2 inhibitor ARN-3261 alone (IA) and in combination with weekly paclitaxel (IB) measuring pharmacokinetics of ARN-3261 and paclitaxel, predictive (SIK2) and pharmacodynamic (pSIK2 and pHDAC5) biomarkers, as well as levels of polyploidy, pAKT, survivin and biomarkers for apoptosis; 2) to determine whether the SIK2 inhibitor ARN-3261 enhances response to carboplatin/paclitaxel and carboplatin/liposomal doxorubicin in ovarian cancer cell line-derived and patient-derived xenografts; and 3) to identify targets that produce synthetic lethality in ovarian cancer cells treated with SIK2 inhibitors.